Theileria parva-Transformed T Cells Show Enhanced Resistance to Fas/Fas Ligand-Induced Apoptosis

Abstract

Lymphocyte homeostasis is regulated by mechanisms that control lymphocyte proliferation and apoptosis. Activation-induced cell death is mediated by the expression of death ligands and receptors, which, when triggered, activate an apoptotic cascade. Bovine T cells transformed by the intracellular parasite Theileria parva proliferate in an uncontrolled manner and undergo clonal expansion. They constitutively express the death receptor Fas and its ligand, FasL but do not undergo apoptosis. Upon elimination of the parasite from the host cell by treatment with a theilericidal drug, cells become increasingly sensitive to Fas/FasL-induced apoptosis. In normal T cells, the sensitivity to death receptor killing is regulated by specific inhibitor proteins. We found that anti-apoptotic proteins such as cellular (c)-FLIP, which functions as a catalytically inactive form of caspase-8, and X-chromosome-linked inhibitor of apoptosis protein (IAP) as well as c-IAP, which can block downstream executioner caspases, are constitutively expressed in T. parva-transformed T cells. Expression of these proteins is rapidly down-regulated upon parasite elimination. Antiapoptotic proteins of the Bcl-2 family such as Bcl-2 and Bcl-xL are also expressed but, in contrast to c-FLIP, c-IAP, and X-chromosome-linked IAP, do not appear to be tightly regulated by the presence of the parasite. Finally, we show that, in contrast to the situation in tumor cells, the phosphoinositide 3-kinase/Akt pathway is not essential for c-FLIP expression. Our findings indicate that by inducing the expression of antiapoptotic proteins, T. parva allows the host cell to escape destruction by homeostatic mechanisms that would normally be activated to limit the continuous expansion of a T cell population.

Akt inhibition upregulates FasL, downregulates c-FLIPs and induces caspase-8-dependent cell death in Jurkat T lymphocytes

“…Additionally, it should be noted that the requirement of Akt kinase to support c-FLIP s expression may differ depending on the cell type and its activation state. Recently, Kuenzi et al 43 reported that the PI3K inhibitor LY did not downregulate c-FLIP expression in T cells of bovine origin transformed by Theileria parva parasite infection. These results indicate that Akt kinase can regulate Fas-mediated apoptotic signaling in a differential and cell/signaling context-dependent manner.…”

c-Myc activation by Theileria parasites promotes survival of infected B-lymphocytes

“…In contrast, survival of a significant proportion of infected lymphocytes has been ascribed to the parasite permanently activating the NF-kB signalling pathway, probably through physical association with the IkB-signalosome (Heussler et al, 2002), but the actual mechanism of NF-kB-mediated protection remains elusive. Nonetheless, it is clear that the presence of live intracellular parasites leads to constitutive induction of a number of antiapoptotic proteins (Kuenzi et al, 2003) and a decrease in proapoptotic ones (Guergnon et al, 2003a), suggesting that Theileria somehow alters the balance of these key regulators in favour of survival. Infected lymphocytes also use autocrine loops to augment their proliferation and we have shown that a TNF-autocrine loop contributes to NF-kB activation, yet inhibition of TNF did not provoke apoptosis (Guergnon et al, 2003a).…”

Promotion of cell death or neurite outgrowth in PC-12 and N2a cells by the fungal alkaloid militarinone A depends on basal expression of p53

“…Immunoblot analysis was performed according to standard procedures essentially as described [30]. Nitrocellulose membranes were blocked and incubated overnight at 4°C with specific primary Ab diluted in blocking buffer (5% BSA in TBS-Tween): anti-PKB 1:1000; anti-Phospho-PKB (Ser473) 1:1000; anti-ERK1/2 1:2000; anti-Phospho-ERK1/2 (Thr202/Tyr204) 1:1000; anti-AIF 1:2000; antiPhospho-p53 (Ser15) 1:2000 (all rabbit polyclonal); mouse monoclonal anti-p53 1:1000 (all Cell Signaling Technology, Beverly, MA, USA); goat polyclonal anti-Actin 1:4000 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).…”

“…To detect caspase activation, staining with FITC-valylalanyl-aspartic acid fluoromethyl ketone (FITC-VAD-fmk; Promega) was carried out as described [30]. Annexin V staining of exposed membrane phosphatidylserine was carried out using the Annexin V assay kit (Roche Diagnostics, Rotkreuz, Switzerland) following the manufacturer's protocol.…”